Endoscopic ultrasound-guided fine-needle aspiration biopsy coupled with KRAS mutation assay to distinguish pancreatic cancer from pseudotumoral chronic pancreatitis.

BACKGROUND AND STUDY AIMS: Differential diagnosis between pancreatic adenocarcinoma (PADC) and pseudotumoral forms of chronic pancreatitis remains difficult. Mutation of KRAS oncogene is present in 75% to 95% of PADC. This study aimed to evaluate whether the combined analysis of KRAS mutation with cytopathological findings from endoscopic ultrasound-guided fine-needle aspiration biopsy (EUS-FNAB) might improve discrimination between PADC and chronic pancreatitis. PATIENTS AND METHODS: This prospective multicenter study included 178 patients with solid pancreatic masses (men 104, women 74; mean age 64.5 years). Cytopathological examination and KRAS mutation analysis (codon-12 and codon-13, restriction fragment length polymorphism [RFLP] and direct sequencing) were performed on EUS-FNAB material. Final diagnoses were obtained on EUS-FNAB analysis and/or a second biopsy and/or clinical follow-up and/or surgery: PADC, n = 129; chronic pancreatitis, n = 27; other pancreatic neoplasms, n = 16; and benign lesions, n = 6. RESULTS: KRAS status analysis was successful in all EUS-FNAB samples. Codon-12 KRAS point mutation was found in 66% of PADC samples. No case of chronic pancreatitis displayed KRAS mutation. Sensitivity, specificity, positive and negative predictive values, and overall accuracy of cytopathology alone for diagnosis of PADC versus chronic pancreatitis were 83%, 100%, 100%, 56% and 86%, respectively. When KRAS mutation analysis was combined with cytopathology, these values reached 88%, 100%, 100%, 63% and 90% respectively. CONCLUSION: Although the value of KRAS analysis in addition to EUS-FNAB is limited for distinguishing pancreatic mass lesions, when chronic pancreatitis presented as a pseudotumor a negative finding (wild-type KRAS), was useful in strongly suggesting a benign lesion.

Replication-deficient rSV40 mediate pancreatic gene transfer and long-term inhibition of tumor growth.

Pancreatic cancer is one of the most aggressive and devastating human malignancies. There is an urgent need for more effective therapy for patients with advanced disease. In this context, genetic therapy potentially represents a rational new approach to treating pancreatic cancer, which could provide an adjunct to conventional options. Because of the promise of recombinant SV40 vectors, we tested their ability to deliver a transgene, and to target a transcript, so as to inhibit pancreatic tumors growth in vivo. BxPC3 and Capan-1 cells were efficiently transduced using SV40 vectors without selection, as compared to synthetic vectors PEI. SV40 vectors were as efficient as adenoviral vectors, and provided long-term transgene expression. Next, we devised a SV40-derived, targeted gene therapy approach of pancreatic cancer, by combining hTR tumor-specific promoter with sst2 somatostatin receptor tumor-suppressor gene. In vitro cell proliferation was strongly impaired following administration of SV(hTR-sst2). SV40-derived sst2-mediated antiproliferative effect was dependent on the local production of somatostatin. In vivo, intratumoral gene transfer of sst2 using rSV40 vectors resulted in a marked inhibition of Capan-1 tumor progression, and proliferation. These results represent the initial steps toward a novel approach to the gene therapy of pancreatic cancer using SV40 as a vector.

Transcription of SST2 somatostatin receptor gene in human pancreatic cancer cells is altered by single nucleotide promoter polymorphism.

BACKGROUND & AIMS: The somatostatin receptor SST2 mediates the antiproliferative effect of stable somatostatin analogues. SST2 gene expression is lost in most human pancreatic carcinomas. We investigated the mechanisms that could be involved in this defect. METHODS: SST2 gene structure was investigated by sequencing and restriction fragment length polymorphism. Characterization of the polymorphism was performed by electrophoretic mobility shift, cross-linking, and transcription assays. RESULTS: No major deletion of the SST2 coding sequence was found in pancreatic carcinoma specimens, but 2 point mutations were frequently detected in the promoter sequence at positions -83 (A-->G) and -57 (C-->G) from the major transcription initiation site. These mutations were present in pancreatic cancer but also in normal pancreatic tissues or leukocytes and thus correspond to a genetic polymorphism. In the 2 human pancreatic cancer cell lines MiaPaCa-2 and AsPC-1, the naturally occurring mutation -57G had no effect on transcription of SST2 gene, whereas -83G mutation reduced it by 60%-70%. We showed that the -83G mutation creates a specific binding site for the nuclear factor I. Cotransfection experiments showed that the nuclear factor I-A1.1 isoform was responsible for SST2 promoter repression. CONCLUSIONS: The -83G polymorphism identified on human SST2 gene promoter is responsible for the specific fixation of nuclear factor I and repression of SST2 transcription in human pancreatic cancer cells. However, its contribution to pancreatic tumorigenesis remains unknown.

Arginine 197 of the cholecystokinin-A receptor binding site interacts with the sulfate of the peptide agonist cholecystokinin.

The knowledge of the binding sites of G protein-coupled cholecystokinin receptors represents important insights that may serve to understand their activation processes and to design or optimize ligands. Our aim was to identify the amino acid of the cholecystokinin-A receptor (CCK-AR) binding site in an interaction with the sulfate of CCK, which is crucial for CCK binding and activity. A three-dimensional model of the [CCK-AR-CCK] complex was built. In this model, Arg197 was the best candidate residue for a ionic interaction with the sulfate of CCK. Arg197 was exchanged for a methionine by site-directed mutagenesis. Wild-type and mutated CCK-AR were transiently expressed in COS-7 cells for pharmacological and functional analysis. The mutated receptor on Arg197 did not bind the agonist radioligand 125I-BH-[Thr, Nle]-CCK-9; however, it bound the nonpeptide antagonist [3H]-SR27,897 as the wild-type receptor. The mutant was approximately 1,470- and 3,200-fold less potent than the wild-type CCK-AR to activate G proteins and to induce inositol phosphate production, respectively. This is consistent with the 500-fold lower potency and 800-fold lower affinity of nonsulfated CCK relative to sulfated CCK on the wild-type receptor. These data, together with those showing that the mutated receptor failed to discriminate nonsulfated and sulfated CCK while it retained other pharmacological features of the CCK-AR, strongly support an interaction between Arg197 of the CCK-AR binding site and the sulfate of CCK. In addition, the mutated CCK-AR resembled the low affinity state of the wild-type CCK-AR, suggesting that Arg197-sulfate interaction regulates conformational changes of the CCK-AR that are required for its physiological activation.

Evidence for a direct interaction between the penultimate aspartic acid of cholecystokinin and histidine 207, located in the second extracellular loop of the cholecystokinin B receptor.

Recently, we reported that the mutation of His(207) to Phe located in the second extracellular loop of the cholecystokinin B receptor strongly affected cholecystokinin (CCK) binding (Silvente-Poirot, S., Escrieut, C., and Wank, S. A. (1998) Mol. Pharmacol. 54, 364-371). To characterize the functional group in CCK that interacts with His(207), we first substituted His(207) to Ala. This mutation decreased the affinity and the potency of CCK to produce total inositol phosphates 302-fold and 456-fold without affecting the expression of the mutant receptor. The screening of L-alanine-modified CCK peptides to bind and activate the wild type and mutant receptors allowed the identification of the interaction of the C-terminal Asp(8) of CCK with His(207). The H207A-CCKBR mutant, unlike the wild type receptor, was insensitive to substitution of Asp(8) of CCK to other amino acid residues. This interaction was further confirmed by mutating His(207) to Asp. The affinity of CCK for the H207D-CCKBR mutant was 100-fold lower than for the H207A-CCKBR mutant, consistent with an electrostatic repulsion between the negative charges of the two interacting aspartic acids. Peptides with neutral amino acids in position eight of CCK reversed this effect and displayed a gain of affinity for the H207D mutant compared with CCK. To date, this is the first report concerning the identification of a direct contact point between the CCKB receptor and CCK.

Gastrin induces phosphorylation of eIF4E binding protein 1 and translation initiation of ornithine decarboxylase mRNA.

Gastrin via its G-protein coupled specific receptor induces transcription of c-fos and c-jun genes through a ras-MAPK pathway. Ornithine Decarboxylase (ODC), a growth regulated proto-oncogene, was chosen to investigate gastrin effects on translation initiation of mRNAs exhibiting a 5'UnTranslated Region (5'UTR) responsible for translation repression in quiescent cells. In AR4-2J tumoral cells, we first demonstrated that gastrin increases ODC mRNA translation. Transient transfections with various CAT chimeric constructs suggested a direct involvement of the 5'UTR in this observation. Translation of this group of mRNAs is enhanced by the availability of the cap-binding protein (eIF4E) that is increased after phosphorylation of its specific binding protein eIF4E-BP1. We found that AR4-2J cells over-expressed eIF4E protein which was not modulated by gastrin treatment. Rapamycin which inhibits 4E-BP1 phosphorylation, completely prevents gastrin-mediated increase of ODC translation indicating that 4E-BP1 could be involved in regulating ODC translation. Implication of 4E-BP1 in mediating gastrin effects is corroborated by the capacity of the ligand to affect 4E-BP1 phosphorylation. These results indicate that gastrin enhances ornithine decarboxylase mRNA translation through a rapamycin sensitive pathway and provide the first evidence in the control of 4E-BP1 phosphorylation after occupancy of a G protein-coupled receptor.

Intraduodenal free fatty acids rather than triglycerides are responsible for the release of CCK in humans.

Exocrine pancreas from different species behaves differently in response to the presence of intact or digested nutrients in the duodenum. A failure of cholecystokinin (CCK) release after a meal has been shown among patients with exocrine pancreatic insufficiency. This abnormality could be restored by the administration of pancreatic extracts, suggesting that digested rather than intact nutrients are responsible for the release of CCK and subsequently gallbladder contraction in humans. The aim of this study was to determine the specific role of different lipidic stimuli in humans. Seven male patients (mean age, 52 years) with pancreatic insufficiency secondary to chronic pancreatitis were selected. Pancreatic insufficiency was considered severe in five of them (lipase output, < 1,000 IU/min) and moderate in another two (lipase output, > 1,000 and < 2,300 IU/min). Plasma CCK (by bioassay), gallbladder contraction (by ultrasound), and enzyme output (chymotrypsin) in response to duodenal administration of either oleic acid as free fatty acids or 20% Intralipid as triglycerides were measured in each patient with at least a 48-h interval between each test. In all these patients with pancreatic insufficiency, duodenal perfusion of free fatty acids generated a more pronounced (91 +/- 11 vs. 49 +/- 21 pM) and faster (15 vs. 30 min) (p < 0.05) CCK release than triglycerides. Furthermore, gallbladder contraction was more efficient when free fatty acids instead of triglycerides were administered in the duodenum (86 +/- 5 vs. 69 +/- 4%) at 10 min (p < 0.05) and (73 +/- 8 vs. 51 +/- 5%) at 15 min (p < 0.03). Among patients with measurable residual pancreatic function, enzyme outputs were shown to be higher during free fatty acid than triglyceride perfusion. In humans, free fatty acids rather than triglycerides, when present in the duodenum, stimulate CCK release and gallbladder contraction. In patients with moderate pancreatic insufficiency this phenomenon may increase residual enzymatic secretion. These results allow us to encourage the development of enzymatic preparations as acid-resistant lipases that cause a fast release of free fatty acids in the duodenum.

AR4-2J cell line coexpresses dihydropyridine and omega-conotoxin sensitive Ca2+ channels.

For the first time, we have demonstrated in AR4-2J cells, an experimental model of azaserine-induced carcinoma in the rat exocrine pancreas, the co-expression of alpha 1 subunit of dihydropyridine-sensitive Ca2+ channel and the alpha 1 sub-unit of omega-conotoxin-sensitive Ca2+ channel RNA messengers which share homologous sequences with, respectively, rbC II and rbB I sub-types described in the rat brain. These two types of voltage-dependent Ca2+ channels which are functionally expressed, emphasize the acquisition during carcinogenesis of neuroendocrine features of AR4-2J cells. Additionally, using antisense phosphorothioate oligodeoxynucleotide, we demonstrated clearly the involvement of dihydropyridine-sensitive Ca2+ channels in the control of AR4-2J cell proliferation.

Autocrine stimulation of AR4-2J rat pancreatic tumor cell growth by glycine-extended gastrin.

Glycine-extended gastrin (gastrin-Gly) stimulates proliferation of AR4-2J pancreatic tumor cell line through a specific receptor, different from the gastrin-cholecystokinin B receptor. Our purpose was to determine whether AR4-2J cells produced gastrin-Gly and then whether the peptide was involved in an autocrine loop. First, proliferation of AR4-2J cells in serum-free medium was inhibited by a gastrin anti-sense oligodeoxynucleotide phosphorothioate and by antibodies specific for gastrin-Gly. In contrast, antibodies specific for alpha-amidated gastrin were without effect. By using RT-PCR, we have shown that AR4-2J cells expressed gastrin mRNA. The presence of gastrin-Gly, but not alpha-amidated gastrin, in serum-free media was detected by radioimmunoanalysis. Gel chromatography revealed that the predominant molecular forms secreted were glycine-extended gastrin-34 and gastrin- 17. Furthermore, epidermal growth factor (EGF), a stimulator of gastrin gene transcription, modulates gastrin-Gly secretion by AR4-2J. These data together suggest that gastrin-Gly is an autocrine growth factor for AR4-2J cells and that it participates with EGF in the regulation of AR4-2J-cell proliferation.

Relief of ornithine decarboxylase messenger RNA translational repression induced by alternative splicing of its 5' untranslated region.

The ornithine decarboxylase enzyme (ODC) is the key regulator of polyamine synthesis and is a member of the cellular proto-oncogene family. Its expression becomes constitutively activated by carcinogens, viruses, and oncogenes. ODC mRNA has a long 5' untranslated region that could be important in the regulation of enzyme levels by affecting translation. To test this hypothesis, we have determined the role of this region on the constitutive ODC hyperexpression measured in AR4-2J cells, an azaserine-induced, tumor-derived pancreatic acinar cell line. Construction of expression vectors in which ODC 5' leader sequence was placed flanking the chloramphenicol acetyltransferase reporter gene allowed us to identify three AR4-2J specific, different alternatively spliced ODC 5' leaders. The 5' ends of exons 2 and 3 were lengthened by 17 and 13 bases, respectively. Translation performed in a cell-free system as well as in COS7 transient transfection experiments demonstrated that AR4-2J isoforms induce a strong increase in the rate of translation. These results provide evidence that alternative splicing observed in tumoral cells, coupled with translation regulation, relieves the translation repression mediated by the long and structured 5' untranslated region of the ODC proto-oncogene.

Identification of K-ras mutations in pancreatic juice in the early diagnosis of pancreatic cancer.

OBJECTIVE: To develop an early diagnostic test for pancreatic cancer based on the identification of K-ras mutations in pure pancreatic juice collected during endoscopic retrograde pancreatography. DESIGN: Prospective study with masked comparison. The standard criteria for the diagnosis of pancreatic cancer were pancreatography or surgery (or both) and histopathology, with follow-up ranging from 6 to 40 months. SETTING: Referral center. PATIENTS: 24 patients with no pancreatic disease (group 1); 29 patients with nontumoral pancreatic disease (group 2); and 22 patients with pancreatic tumor (group 3). Endoscopic ductal aspiration of cells or brush cytology was done on patients having endoscopic retrograde pancreatography for diagnostic or therapeutic reasons. MAIN OUTCOME MEASURE: Confirmation of mutation rates in patients with pancreatic cancer. RESULTS: K-ras gene analysis was done by polymerase chain reaction-mediated restriction fragment length polymorphism analysis and direct sequencing. All patients from groups 1 and 2 (n = 53) had a normal sequence for the K-ras 12th codon (group 1, 0% [95% CI, 0% to 14%]; group 2, 0% [CI, 0% to 12%]). Mutations were seen in 17 of the 22 patients in group 3 (77% [CI, 55% to 92%]). Two of the 17 had no evidence of pancreatic cancer when K-ras was first studied. One had chronic abdominal pain and the other presented with acute pancreatitis. Both were initially free of any pancreatic mass, but they developed tumors 18 and 40 months, respectively, after the K-ras mutations were identified. CONCLUSION: Identification of K-ras mutations in samples of pancreatic juice may be useful in differentiating between pancreatic cancer and noncancerous pancreatic diseases. K-ras mutation can precede clinical evidence of pancreatic cancer, but the clinical implications of this finding need further study.

Photoaffinity labeling of rat pancreatic cholecystokinin type A receptor antagonist binding sites demonstrates the presence of a truncated cholecystokinin type A receptor.

During the past few years, several antagonist ligands for cholecystokinin (CCK) receptors have been discovered, but the mechanism of action of these candidate drugs, as well as the nature of their molecular targets, remains poorly documented. In a previous study, we developed a new antagonist radioligand, 125I-Bolton-Hunter-labeled JMV-179, for the CCK-A receptor (CCK-AR), to analyze CCK antagonist binding sites in pancreatic plasma membranes. We found that 125I-Bolton-Hunter-labeled JMV-179 identified 4 times as many sites as did an agonist radioligand, although agonists were able to interact competitively with the entire population of antagonist sites. In the present work, using biochemical approaches we have identified and characterized CCK antagonist binding sites in pancreatic plasma membranes. We synthesized the photoactivable antagonist probe 125I-azidosalicyclic acid (ASA)-JMV-179. The binding of 125I-ASA-JMV-179 to plasma membranes was inhibited by JMV-179 (IC50, 6 +/- 2 nM), by (Thr28, Ahx31)-CCK-25-33 (IC50, 1.2 +/- 0.5 nM), and by the nonpeptide CCK-AR antagonist L-364,718 (IC50, 2 +/- 1 nM). Photoaffinity labeling using pancreatic membranes or acini demonstrated that 125I-ASA-JMV-179 detected a new 47-50-kDa protein in addition to the 85-100-kDa CCK-AR. The 47-50-kDa protein was not directly detected by a photoactivable agonist, but agonists could inhibit its covalent labeling by 125I-ASA-JMV-179 (IC50 for (Thr28,Ahx31)-CCK-25-33, 15 nM). In competition assays using nonsolubilized or solubilized membranes, this protein displayed binding features of the CCK-AR and was retained on immobilized wheat germ agglutinin, as was the CCK-AR. To further characterize the 47-50-kDa protein, deglycosylation and protease digestions were performed, and the digestion products were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Protease digestions of both the CCK-AR and the 47-50-kDa protein yielded identical labeled fragments, demonstrating a structural relationship between the two proteins. The CCK-AR, which has three potential sites for N-glycosylation on the amino-terminal extracellular domain and one on the second extracytoplasmic loop, was deglycosylated to a 42-kDa peptide. The 47-50-kDa protein was deglycosylated to a 35-kDa peptide. These data, and the localization of the labeled fragments in the amino acid sequence of the receptor, suggest that the 47-50-kDa protein represents a CCK-AR lacking its amino-terminal extracellular domain.(ABSTRACT TRUNCATED AT 400 WORDS)

Postprandial cholecystokinin secretion in elderly with protein-energy undernutrition.

OBJECTIVE: Malnutrition is currently observed in aged people, and cholecystokinin is an important peripheral satiety signal. The aim of this study was to examine the effect of aging and protein-energy malnutrition on postprandial cholecystokinin (CCK) release. DESIGN: Non-randomized, cross-sectional comparison by age group. SETTING: Gastroenterology section of a teaching hospital. PARTICIPANTS: Twenty-one human volunteers divided into three groups: young healthy subjects (Group 1: mean 29 years, n = 7), aged healthy subjects (Group 2, mean 80 years, n = 7), and aged subjects with an important degree of malnutrition (Group 3, mean 84.6 years, n = 7). INTERVENTION: Each subject ingested a standardized liquid meal after an overnight fast. MAIN OUTCOME MEASURES: Plasma cholecystokinin was measured using a sensitive bioassay before and after the ingestion of the liquid meal. RESULTS: Basal cholecystokinin levels were similar (0.9 to 1 pM equivalent CCK-8) in the three groups. Postprandial levels were significantly increased over basal (P less than 0.05). The maximal cholecystokinin value was lower in Group 1 (3.5 +/- 0.8 pM equivalent CCK-8) and Group 2 (3.3 +/- 0.77 pM equivalent CCK-8) than in Group 3 (8.3 +/- 2 pM equivalent CCK-8) (P less than 0.05). Integrated plasma cholecystokinin was also similar in Group 1 (171 +/- 38 pM.60 min), (P less than 0.05). CONCLUSION: The increase of postprandial maximal levels of cholecystokinin is more related to malnutrition than to aging.

Gastric lipase in alcoholic pancreatitis. Comparison of secretive profiles following pentagastrin stimulation in normal adults and patients with pancreatic insufficiency.

The aims of this study were to evaluate the amount of gastric lipase secreted by the stomach in normal adults and to elucidate a possible adaptative secretion of this enzyme in response to pancreatic insufficiency secondary to alcoholic chronic pancreatitis. Forty-one subjects underwent a gastric intubation. Pentagastrin (6 micrograms.kg-1.h-1 IV) significantly increased gastric lipase concentration and output. Stimulated gastric lipase output in seven normal subjects was 12,598 +/- 2036 U/h (by using tributyrin as substrate). Outputs where higher (P less than 0.02) in 17 patients with pancreatic insufficiency who were not drinking alcohol, but were not significantly different in nine patients who continued to drink (20,413 +/- 1778 U/h and 21,953 +/- 4973 U/h, respectively). On the other hand, high gastric lipase outputs were found in eight patients with duodenal ulcers and no evidence of pancreatic dysfunction (23,180 +/- 262 U/h). The time required to reach maximal lipase output (peak output) following pentagastrin stimulation was the same in all groups (approximately 38 minutes) except for the group of patients with pancreatic insufficiency who did not drink alcohol, in whom it was significantly reduced (approximately 26.5 minutes). Secretory patterns of gastric lipase and pepsin were closely comparable. Gastric lipase secretion could be increased in several clinical conditions and particularly in patients with pancreatic insufficiency caused by alcoholic chronic pancreatitis who have been abstinent for a long time.

Protective effect of misoprostol, a synthetic prostaglandin E1 analog, on cerulein-induced acute pancreatitis in rats.

This study was performed to assess the effects of misoprostol, a synthetic prostaglandin E1 analog, on cerulein-induced pancreatitis. Per group of 10 each, male Wistar rats received either cerulein (2.5 micrograms/kg/h subcutaneously), cerulein and misoprostol (500 micrograms/kg intraperitoneally at 0 and 4 h), or saline. Rats were killed 6 h after the first injection. Misoprostol treatment significantly reduced interstitial edema and acinar cell lesions induced by hyperstimulation. Pancreatic amylase and chymotrypsin contents were increased by cerulein and returned towards control levels in the misoprostol-treated group. The lysosomal volume density and the pancreatic beta-D-glucuronidase activity were significantly increased after hyperstimulation. The two parameters were significantly reduced by misoprostol. A protective effect of misoprostol against lesions induced by cerulein hyperstimulation would be a consequence of a lysosomal stabilizating effect.

Protective effect of misoprostol, a synthetic prostaglandin E1 analog, on experimental pancreatitis induced by pancreatic duct ligation in rat.

This study was performed to assess the effects of misoprostol (M), a synthetic prostaglandin E1 analog, on experimental pancreatitis in rat. Pancreatitis was induced by ligation of the main pancreatic duct of 3-month-old male Wistar rats. Pancreatic lesions were observed at 6, 12, 24, 48, and 96 h after pancreatic duct ligation (PDL). A time of 48 h was chosen to evaluate M treatment. M was injected intraperitoneally (500 micrograms/kg every 4 h) between time 0 and 48 h after PDL. Stereological analysis was performed on light and electron microscopy. Total pancreatic amylase and chymotrypsin concentrations were determined. Four groups of five rats were studied: sham operated (SO), M without PDL (PG), duct ligation without M (DL), and duct ligation with M (DLPG). Edema, dedifferentiation of pancreatic acinar cells, and heterogeneous distribution of zymogen granule diameters observed after PDL were significantly decreased by M in the DLPG group. Enzyme concentrations were also decreased by M in the DLPG group. Enzyme concentrations were also decreased by M both in normal (PG) and duct ligated rats (DL). M has protective effects against pancreatic lesions induced by PDL. In this model, the protective effect of M may be due to a blockade of the autodigestive secretions of the pancreatic acinar cells.

Exocrine pancreatic secretion in the elderly.

In order to evaluate impairment of exocrine pancreatic function during aging, 27 subjects (mean age: 36 years +/- 7.8) and 28 subjects (mean age: 72 years +/- 3.2), with no clinical or radiological evidence of digestive disease, were selected. Duodenal aspirates over a 60 min period were obtained during continuous IV infusion of secretin (0.5 U/kg/h) and caerulein (75 ng/kg/h). Bicarbonate, lipase, chymotrypsin amylase concentrations and output were measured. Bicarbonate, lipase, chymotrypsin concentrations in the aged group were significantly reduced by 17%, 15% and 23% respectively (P less than 0.05) as compared with those in the young group. In addition, a significant reduction of approximately 45% in bicarbonate and enzyme output levels was observed. This study provides strong evidence for a marked functional involution of the exocrine pancreatic secretion during aging. The potential consequences of this phenomenon on the nutritional status in the elderly are discussed.

[Aging of the pancreas. Its implication in malnutrition states in elderly persons]. / Vieillissement du pancréas. Son implication dans les états de dénutrition chez les personnes âgées.

The influence of ageing on exocrine pancreatic function was investigated in rats and in men. Young rats (3-months old, 150-200 g) and old rats (24-month old, 400-500 g) were killed after fasting overnight. Small pancreatic fragments were removed and kept for stereological analysis both in light and electron microscopy; in addition, levels of tritium-labelled leucine uptake and protein synthesis were measured by quantitative histoautoradiography on isolated acini in vitro. The human study was conducted retrospectively in 27 adults (mean age 36 +/- 1.5 years) and in 28 elderly subjects (mean age 72 +/- 0.6 years) with no clinical or radiological evidence of pancreatitis. Duodenal aspirates were taken over a 90 min period under secretin (0.5 U/kg.h) and cerulein (75 U/kg.h) infusion for comparisons of bicarbonate, lipase, chymotrypsin and amylase concentrations and outputs in the two groups. Elderly people were found to have significant and parallel decreases in bicarbonate, lipase and amylase output as compared to younger subjects (-40 per cent, P less than 0.001). The pancreatic deficiency was confirmed in rats by a significant decrease in zymogen volume density and zymogen diameter and a defect of protein synthesis with significant slowing down (-70 per cent, P less than 0.001) of newly synthesized protein transfer to the Golgian zone of acinar cells. Signs of exocrine parenchyma dystrophy (pancreatitis?) were also observed.

[Comparison of fungal lipase and pancreatic lipase in exocrine pancreatic insufficiency in man. Study of their in vitro properties and intraduodenal bioavailability]. / Comparaison d'une lipase fungique et d'une lipase pancréatique au cours de l'insuffisance pancréatique exocrine chez l'homme. Etude de leur propriété in vitro et leur biodisponibilité intraduodénale.

Assuming that acidic degradation of lipase was the major cause of failure for the correction of steatorrhea by pancreatic extracts, we compared the in vitro and in vivo activities of a fungal lipase (FL) (Rhizopus arrhizus) with classical porcine pancreatic extract (Eurobiol). The choice of FL was determined by its two optimum pH (3.5 and 7.4). Five factors known to modify lipase activity were tested: pH, biliary acids colipase, trypsin and albumin. Bioavailability was measured by using a double intubation method in 13 patients with severe pancreatic insufficiency. Each enzymatic preparation was given during a test meal in a randomized and cross-over fashion. Results of the in vitro study showed that FL differed from pancreatic lipase by the following properties: better resistance in acidic solution, inhibition by biliary salts, absence of effect of colipase and rapid degradation by trypsin. In vivo the percentage of lipase activity recovered was 14.2 +/- 10.6 p. 100 for FL and 56 +/- 50 p. 100 for the classical pancreatic preparation. Compared with placebo significant differences in the recovery rate of lipolytic activity were observed with the pancreatic preparation only and started at the 40th min after the end of the test meal. These results showed that lack of degradation in acidic milieu is not the only valuable criterion for the choice of an efficient lipase preparation. The role of other potential factors such as gastric emptying as well as proteolytic degradation of the enzyme should be considered as well.

The Ebner glands: a pancreatic-like gland secreting an acid lipase. Secretory regulation in vitro.

The lingual serous glands of rat tongue, the Ebner glands, secrete a potent acid lipase that acts in the stomach where it initiates the digestion of dietary fat. The factors affecting its secretion were studied 'in vitro' on Ebner slices. The lipolytic activity was measured in the incubation medium using tributyrine as substrate and by titration at pH: 5.4. Cholecystokinin (10(-9) M) and carbachol (10(-5) M) efficiently stimulated lipase secretion (3 fold over basal rate). The parasympathetic agent triggered secretion involving a calcium dependent system. Atropin (10(-4) M) blocked this cholinergic effect by about 40%. Lipase secretion was stimulated by epinephrine and isoproterenol. Propranolol (beta-antagonist) inhibited the adrenergic stimulation, while phentolamine was ineffective. The inhibitory effect of the selective beta 1-antagonist (Betaxolol) and the lack of effect of the selective beta 2-antagonist (ICI 118551) suggest the participation of beta 1-adrenoceptors in the secretion mechanism.